The invention relates to repairing and regenerating damaged tissue in a mammal (e.g., a human patient). Such damage may result from an existing autoimmune disease, or may be the result of a non-autoimmune insult. I have previously shown that eliminating autoimmune cells and re-educating the immune system are important components of an effective treatment of an autoimmune disease (described in U.S. patent application Ser. Nos. 10/358,664, 09/521,064, 09/768,769, and Ryu et al., Journal of Clinical Investigations, 108: 31-33, 2001, which are hereby incorporated by reference herein). While an autoimmune disease may be successfully treated, the individual may nonetheless have significant tissue damage as a result of the prior autoimmune attack.
Many tissues have an innate ability to repair themselves once the damage causing insult is eliminated, but this ability to repair damage decreases in correlation with the duration of the insult. For example, the regenerative capacity of endogenous pancreatic islets is virtually eliminated in long-term Type I diabetics, i.e., patients who have had the disease for more than 15 years. In cases where the endogenous tissue has lost its regenerative capacity, the damage may be repaired by providing exogenous tissue to the individual, for example, by a transplant. A promising treatment for diabetes, islet transplantation, has been the subject of human clinical trials for over ten years. While there have been many successes with islet transplantation in animals, these have occurred where the animals are diabetic due to chemical treatment, rather than natural disease. The only substantiated peer reviewed studies using non-barrier and non-toxic methods and showing success with islet transplants in naturally diabetic mice use isogeneic (self) islets. The isogeneic islets were transplanted into non-obese diabetic (NOD) mice with active diabetes, which were pre-treated with TNF-alpha (tumor necrosis factor-alpha); BCG (Bacillus Clamette-Guerin, an attenuated strain of mycobacterium bovis); or CFA (Complete Freund's Adjuvant), which is an inducer of TNF-alpha (Rabinovitch et al., J. Immunol. 159: 6298-6303, 1997). This approach is not clinically applicable primarily because syngeneic islets are not available. Furthermore, existing cell replacement strategies have not prevented end-stage diseases or permanently reversed insulitis. In the allograft setting of islet transplantation, grafts are eventually rejected, even with immunosuppression. Furthermore, diabetic host treatments such as body irradiation and bone marrow transplantation are unacceptably toxic, rendering the short-term alternative of insulin therapy more attractive.
Recently, islet transplantation has achieved limited success in clinical trials, with type 1 diabetic patients having a sustained return to normoglycemia over a 6 month period. These results have been obtained with continuous, and sometimes toxic, drug therapy, often in the setting of a simultaneous life-saving renal transplant. However, these moderately successful islet transplants show failures after about one year, speculated to be due in part to the drug therapy itself inducing insulin resistance. The earlier failure of islet transplants in type 1 diabetics, compared to non-diabetic patients receiving islet transplants (such as in cancer patients who have had their pancreas removed), raises the concern that immunosuppressive therapy shows greater efficacy for graft rejection over autoimmunity prevention. Lending credence to these concerns is the observation of the inefficiency of immunosuppression therapy for the prevention of graft rejection of allogenic or xenogeneic islet transplants in animal studies using non-obese diabetic (NOD) mice.
I have previously described a transplantation method to introduce allogeneic and xenogeneic tissues into non-immunosuppressed hosts in which the cells are modified such that the donor antigens are disguised from the host's immune system (U.S. Pat. No. 5,283,058, which is hereby incorporated by reference). Generally, masked islets or transgenic islets with ablated MHC class I molecules are only partially protected from recurrent autoimmunity in NOD mice (Markmann et al., Transplantation 54: 1085-89, 1992). It has also been shown that a brief two-component therapy is able both to reestablish self-tolerance and to eliminate selectively the pathological memory T cells of NOD mice by the induction of apoptosis (Ryu et al., Journal of Clinical Investigations, 108: 31-33, 2001). Simultaneous treatment of severely diabetic animals with TNF-α (or an inducer of endogenous TNF-α production) and with splenocytes partially or fully matched with regard to MHC class I antigens (to reselect pathogenic naïve T cells) thus results in permanent reversal of established diabetes. This “cure” is accompanied by the reappearance of insulin-secreting islets in the pancreas of treated animals that are able to control blood glucose concentration in a manner indistinguishable from that apparent in normal mice.
The existence of pluripotent stem cells in the bone marrow of adult mammals has been well documented. The existence of pluripotent cells that reside outside the bone marrow has also been demonstrated (Kuehnle and Goodell, Br. Med. J. 325: 372-6; 2002; Rosenthal, New Eng. J. Med. 349: 267-74; 2003). Studies in both mice and in humans have shown that the introduction of MHC-matched bone marrow cells into irradiated hosts results both in repopulation of the host bone marrow as well as rare examples of donor engraftment of host parenchymal organs, including the liver, brain, muscle, and heart, with scattered tissue-specific cells. Such engraftment is typically neither robust nor durable, however. In culture, pluripotent cells are also able to differentiate into mesoderm, neuroectoderm, and endoderm (Jiang et al., Nature 418: 41, 2002). Presumably, the nonlymphoid cells of donor origin in the transplantation studies are the product of transdifferentiation, the conversion of one adult cell type to another. Furthermore, cultured adult bone marrow stem cells may fuse, albeit at a low frequency, with co-cultured embryonic cells. More robust fusion events occur in the remaining liver tissue in a mouse model of liver damage after total host body irradiation and transplantation of bone marrow cells. However, such fusion generates cells with marked chromosomal abnormalities and does not represent transdifferentiation or developmental plasticity. Concerns have also been raised about the functionality or malignant potential of some cultured pluripotent cells or their in vivo fusion derivatives if the stem cells are to be used for therapeutic purposes in humans.
A need exists for methods of regenerating damaged tissue using adult pluripotent cells. Desirably, the pluripotent cells need little or no damaging pre-treatment (such as, for example, irradiation or chemical treatment), or are from an endogenous source and are induced or stimulated. Ideally, the regeneration methods would not only be applicable to tissue damage that results from autoimmune attack, but also to non-autoimmune induced damage.